Abstract

Cockayne syndrome is a neurodegenerative accelerated aging disorder caused by mutations in the CSA or CSB genes. Although the pathogenesis of Cockayne syndrome has remained elusive, recent work implicates mitochondrial dysfunction in the disease progression. Here, we present evidence that loss of CSA or CSB in a neuroblastoma cell line converges on mitochondrial dysfunction caused by defects in ribosomal DNA transcription and activation of the DNA damage sensor poly-ADP ribose polymerase 1 (PARP1). Indeed, inhibition of ribosomal DNA transcription leads to mitochondrial dysfunction in a number of cell lines. Furthermore, machine-learning algorithms predict that diseases with defects in ribosomal DNA (rDNA) transcription have mitochondrial dysfunction, and, accordingly, this is found when factors involved in rDNA transcription are knocked down. Mechanistically, loss of CSA or CSB leads to polymerase stalling at non-B DNA in a neuroblastoma cell line, in particular at G-quadruplex structures, and recombinant CSB can melt G-quadruplex structures. Indeed, stabilization of G-quadruplex structures activates PARP1 and leads to accelerated aging in Caenorhabditis elegans In conclusion, this work supports a role for impaired ribosomal DNA transcription in Cockayne syndrome and suggests that transcription-coupled resolution of secondary structures may be a mechanism to repress spurious activation of a DNA damage response.

Diseases associated with transcriptional defects display mitochondrial phenotypes. Each dot represents a disease and the connecting lines a shared trait between two diseases. The closer the two diseases are, the more features they share. See www.mitodb.com/network.html for details.

No apparent futile transcription cycles or loss of mitochondrial DNA transcription in CSA or CSB knockdown cells. (A) Quantification of nuclear run-on experiments using probes against the 5′ end of three genes (mean ± SEM, n = 2–3). (B) Quantification of nuclear run-on experiments comparing signal from the 5′ end with the 3′ prime ends of genes (mean ± SEM, n = 2–3). (C) Loss of transcription across genes as a measure of the slope of normalized coverage. (D) Fold change in mitochondrial transcripts normalized to controls. Outer circle shows the map of the mitochondrial DNA.

Transcription in the vicinity of G4-forming sequences in normal young and elderly humans. (A) Transcription in the vicinity of G4 structures on the template and opposite strand in vivo in cells from the iliac crest of healthy young and elderly women. (B) Transcriptional pausing as a function of transcriptional activity (overall coverage) on the template or opposite strand in vivo in cells from the iliac crest of healthy young and elderly women.